Biomedical subjects

I Moro

Publications and source records attributed to I Moro.

At least 19 recordsLinked to original sources

PubMed2005

Modulations of the thylakoid system in snow xanthophycean alga cultured in the dark for two months: comparison between microspectrofluorimetric responses and morphological aspects.

The response of the plastid was studied, with a special emphasis on thylakoid structure and function, in a snow filamentous xanthophycean alga (Xanthonema sp.) incubated in darkness for two months. Microspectrofluorimetric analyses were performed on single living cells to study the variations in the assembly of the chlorophyll-protein complexes of photosystem II, in comparison with cells grown in light. In parallel, changes in micro- and submicroscopic plastid morphology and in photosynthetic pigment content were monitored. Throughout the experiment, the lamellar architecture of thylakoids in the alga was relatively well preserved, whereas photosystem II underwent disassembly and degradation triggered by prolonged darkness. Conversely, the light-harvesting complex of photosystem II proved to be relatively stable for long periods in darkness. Moreover, a role of the peripheral antennae in determining thylakoid arrangement in xanthophycean algae is implied. Although the responses observed in Xanthonema sp. can be considered in terms of acclimation to darkness, the progressive destabilisation of the light-harvesting complex of photosystem II testifies to incipient ageing of the cells after 35 days.

Eukaryota↗

PubMed2004

Active synthesis of mouse polymeric immunoglobulin receptor in the epithelial cells of the distal urinary tubule in kidney.

The tissue distribution of mouse polymeric immunoglobulin receptor (pIgR) has been demonstrated. By Northern blot hybridization, pIgR mRNA expression was detected in liver, intestine, stomach, lung and kidney. A weak expression was also detected in thymus by reverse transcriptase-polymerase chain reaction. The pIgR expression in kidney was further studied and confirmed that pIgR protein was actively synthesized in the epithelial cells of distal urinary tubule and of Henle's loop. Immunoelectron microscopical analysis showed the accumulation of pIgR-containing vesicles in the apical portion of distal urinary tubule epithelial cells.

Animals↗

PubMed2004

Retinoic acid enhances the gene expression of human polymeric immunoglobulin receptor (pIgR) by TNF-alpha.

In this study, the detailed mechanisms for the effects of vitamin A on the expression of polymeric immunoglobulin receptor (pIgR) were examined. Expression of the pIgR by tumour necrosis factor (TNF-alpha) was enhanced by the addition of all-trans retinoic acid (ATRA) or 9-cis retinoic acid (9CRA). This enhancement was mediated mainly by RARalpha, and regulated at the transcriptional level. Transcription factor nuclear factor-kappaB (NF-kappaB) binding and activation were not influenced by addition of ATRA. These data imply that RA, in combination with TNF-alpha, could up-regulate the expression of pIgR. In addition, we hypothesize that up-regulation of pIgR by RA is controlled through the RAR-dependent signalling pathway and that it plays a role in enhancement of mucosal immunity.

Blotting, Northern↗

PubMed2004

Dietary fructooligosaccharides up-regulate immunoglobulin A response and polymeric immunoglobulin receptor expression in intestines of infant mice.

We examined whether or not dietary fructooligosaccharides (FOS) in infancy can have a beneficial effect on the mucosal immune system. Newborn BALB/c mice, accompanied by their dams until 21 days of age, were fed either a control diet based on casein [FOS- diet group] or a FOS- diet supplemented with 5% (w/w) FOS [FOS+ diet group]. Total IgA levels in tissue extracts from the intestines of mice in the FOS+ diet group at 38 days of age were about twofold higher (P < 0.05) than those in the FOS- diet group in the jejunum, ileum and colon. Ileal and colonic polymeric immunoglobulin receptor (pIgR) expression in the FOS+ diet group at 36 days of age was 1.5-fold higher than in the FOS- diet group (P < 0.05). Consistent with these results, the ileal IgA secretion rate of the FOS+ diet group at 37 days of age was twofold higher than that of the FOS- diet group (P < 0.05). Moreover, the percentage of B220(+)IgA+ cells in Peyer's patches (PP) was significantly higher in the FOS+ diet group than in the FOS- diet group (6.2%versus 4.3%, P < 0.05), suggesting that isotype switching from IgM to IgA in PP B cells might be enhanced in vivo. Taken together, our findings suggest that dietary FOS increases the intestinal IgA response and pIgR expression in the small intestine as well as the colon in infant mice.

Animals↗

PubMed2003

Release of non-glycosylated polymeric immunoglobulin receptor protein.

Using a recombinant vaccinia virus containing the T7 RNA polymerase, we have established a system for the transient expression of human polymeric immunoglobulin receptor (pIgR) in baby hamster kidney cells, a baby hamster-derived fibroblastic cell line. This transfection system resulted in the successful expression of pIgR in these cells, and Western blot analysis showed that human pIgR was expressed as two different molecular weight forms of 92 and 107 kDa. Treatment with endoglycosidase H showed that the difference between these two forms was due to the glycosylation status of the protein. In order to examine the functional role of glycosylation, we treated the transfected cells with tunicamycin, which prevents a core glycosylation step in the endoplasmic reticulum. Non-glycosylated pIgR was released into the culture medium of the transfected cells, albeit with extremely low efficiency. Taking these results together, we conclude that the glycosylation of pIgR may play a positive role in the efficient transport or release of free pIgR.

Animals↗

PubMed2001

Ontogeny of the murine Ig joining chain gene and protein.

We used Northern blot analysis in order to investigate the ontogeny of the murine joining (J)-chain gene. No J-chain expression was detected in embryonic tissues, including liver, spleen and intestine, but an expression of mu heavy chain was detected in foetal liver at day 17. J-chain expression was detected in the spleen at day 9 and in the intestine at day 15 after birth. Western blot analysis was carried out in order to compare the protein levels of J and mu heavy chains in serum from day 8 to day 24 after birth, using antihuman J chain and antimouse mu chain antibodies. Although mu chain protein could be detected in serum from day 8, J-chain protein was detectable only at day 24. These results suggest that the expression of J chain is a later event than the mu chain in the mouse, which thus differs in embryogenesis from humans.

Animals↗

PubMed2001

Evidence for perforin-like activity associated with earthworm leukocytes.

Earthworm (Eisenia fetida) coelomic fluid contains several leukocytes (coelomocytes): basophils, acidophils and neutrophils as well as chloragocytes. Small coelomocytes and coelomocyte lysate are cytotoxic for the tumor cell target K562. The expression of a lytic factor was investigated by immunocytochemistry using light and transmission electron microscopy. A rat-anti-mouse-perforin-mAb labeled mainly small coelomocytes (nearly 20%) as visualized by light microscopy. TEM analysis using immunogold showed a homogenous labeling in the cytoplasm of small coelomocytes. The highest number of immunogold particles was estimated in coelomocytes with many small cytoplasmic granules. Coelomocytes with large lysosomal granules were also labeled but less intensely. No antibody binding was observed for chloragocytes either in light or electron microscopy. This suggests that the perforin-like activity is associated with only one cell type and that chloragocytes are responsible for other lytic activities. MALDI-MS revealed calreticulin usually associated with perforin in mammalian cells that mediate lysis (e.g. NK, CTL). Together, results strongly suggest the presence of putative perforin in earthworms. This in turn supports the hypothesis that perforin is a conserved component important in immune defense during evolution.

Journal Article↗

PubMed2001

Responses to bleaching herbicides by leaf chloroplasts of maize plants grown at different temperatures.

The effects of growth temperature on chloroplast responses to norflurazon and amitrole, two herbicides inhibiting carotenogenesis, at phytoene desaturation and lycopene cyclization, respectively, were studied in leaves of maize plants grown at 20 degrees C and 30 degrees C in light. At the lower temperature both chemicals caused severe photo-oxidative damage to chloroplasts. In organelles of norflurazon-treated leaves neither carotenoids nor chlorophylls were detectable and the thylakoid system was dismantled. In organelles of amitrole-treated leaves lycopene was accumulated, but small quantities of beta-carotene and xanthophylls were also produced. Moreover, some chlorophyll and a few inner membranes still persisted, although these latter were disarranged, lacking essential protein components and devoid of photosynthetic function. The increase in plant growth temperature to 30 degrees C did not change the norflurazon effects on carotenoid synthesis and the photo-oxidative damage suffered by chloroplasts. By contrast, in organelles of amitrole-treated leaves a large increase in photoprotective carotenoid biosynthesis occurred, with a consequent recovery of chlorophyll content, ultrastructural organization and thylakoid composition and functionality. This suggests that thermo-modulated steps could exist in the carotenogenic pathway, between the points inhibited by the two herbicides. Moreover it shows that, unlike C(3) species, C(4) species, such as maize, can express a strong tolerance to herbicides like amitrole, when supplied to plants growing at their optimum temperature conditions.

Chloroplasts↗

PubMed2001

CD14 is expressed and released as soluble CD14 by human intestinal epithelial cells in vitro: lipopolysaccharide activation of epithelial cells revisited.

Human endothelial as well as epithelial cells were shown to respond to lipopolysaccharides (LPSs). However, the expression and release of CD14 by these so-called CD14-negative cells have not been studied in detail. We investigated three human intestinal epithelial cell lines (ECLs), SW-480, HT-29, and Caco-2, for their expression of CD14 and CD11c/CD18 as well as their responsiveness to endotoxins. Fluorescence-activated cell sorter analysis revealed no expression of CD11c/CD18, but there was low expression of membrane-bound CD14 on HT-29, Caco-2, and SW-480 ECLs. Both Western blotting and reverse transcription-PCR confirmed the CD14 positivity of all three intestinal ECLs. No substantial modulation of CD14 expression was achieved after 6, 8, 18, 24, and 48 h of cultivation with 10-fold serial dilutions of LPS ranging from 0.01 ng/ml to 100 microg/ml. Interestingly, soluble CD14 was found in the tissue culture supernatants of all three ECLs. Finally, only HT-29 and SW-480, and not Caco-2, cells responded to LPS exposure (range, 0.01 ng/ml to 100 microg/ml) by interleukin 8 release. Thus, we show that HT-29, SW-480, and Caco-2 human intestinal ECLs express membrane-bound CD14. As Caco-2 cells did not respond to LPS, these cell lines might be an interesting model for studying the receptor complex for LPS. The fact that human intestinal epithelial cells are capable not only of expression but also of release of soluble CD14 may have important implications in vivo, e.g., in shaping the interaction between the mucosal immune system and bacteria in the gut and/or in the pathogenesis of endotoxin shock.

Blotting, Western↗

PubMed2001

Histopathological and immunohistological analyses in IgA deficient lymphoplasia (aly/aly) mouse.

The aly/aly mouse has a severe immunodeficiency, because it lacks peripheral lymph nodes as well as IgA and IgG immunoglobulin synthesis. In the present study, we performed histopathological and immunohistological examinations to clarify histological disorders of various immune organs in these mice. Carbon CH40 injections into the apex of the tongue confirmed the absence of submandibular lymph nodes in aly/aly mice. The thymus had a poorly constructed cortex and medulla, and the number of lymphoid follicles was clearly decreased in the spleen. No IgG- or IgA- producing cells were found in any immune organs, including the mucosal immune sites, though several IgM -producing cells were identified. Other characteristic findings included perivascular lymphocytes accumulation in the salivary glands, lungs, liver and pancreas, which caused tissues damage. These results demonstrated that the various lymphoid tissues disorders and organ-specific lymphocyte infiltration cause immuno-deficiency in the aly/aly mouse.

Animals↗

PubMed2000

Cloning and expression of the chicken immunoglobulin joining (J)-chain cDNA.

The J chain is a component of polymeric immunoglobulin (Ig) molecules and may play an important role in their polymerization and the transport of polymeric Ig across epithelial cells. In this study, the primary structure of the chicken J chain was determined by sequencing cDNA clones. The cDNA had an open reading frame of 476 nucleotides encoding a putative protein of 158 amino acid residues including the signal sequence. The 3' untranslated region consisted of 1216 nucleotides and a poly(A) tail. The deduced amino acid sequence of the chicken J chain had a high degree of homology to that of human, cow, rabbit, mouse, frog, and earthworm, with eight conserved Cys residues identical to the mammalian J chains. Northern blot hybridization performed with total RNA from various chicken tissues revealed high levels of J-chain mRNA expression in spleen, intestine, Harderian gland, and bursa of Fabricius, and low levels in the thymus. The J chain was expressed in the bursa as early as day 15 of embryogenesis. These data indicated that the chicken J-chain gene displays a high degree of homology with that of other species, and is expressed at an early stage of development of the chicken immune system.

Amino Acid Sequence↗

PubMed2000

Role of nuclear factor-kappaB in the expression by tumor necrosis factor-alpha of the human polymeric immunoglobulin receptor (plgR) gene.

We analyzed the mechanism of human polymeric immunoglobulin receptor (pIgR) gene upregulation by tumor necrosis factor (TNF)-alpha. Northern blot analysis showed that the expression of pIgR mRNA was enhanced by TNF-alpha stimulation. This activation was completely inhibited by RNA polymerase or protein synthesis inhibitors, suggesting that the regulation of pIgR gene expression depends on de novo RNA and protein synthesis. Furthermore, the stimulation of pIgR mRNA by TNF-alpha was decreased by pyrrolidinedithiocarbamate and L-1-4'-tosylamino-phenylethyl-chloromethyl ketone, which are known nuclear factor (NF)-kappaB inhibitors. For further analysis of gene regulation, we cloned and sequenced the 1.5-kb 5'-flanking region of the pIgR gene. In the upstream region, we found two NF-kappaB-binding motifs (named kappaB1 and kappaB2 from the 5' region). An electrophoretic mobility shift assay indicated that two components of the NF-kappaB/Re1 family, p50 and p65, bound with higher affinity to the KB2 element than to the kappaB1 element. We also analyzed plgR gene expression using reporter plasmids expressing the firefly luciferase gene. Stimulation by TNF-alpha significantly activated the pIgR gene promoter, as a 775-bp upstream region of the pIgR gene increased luciferase gene expression in cells treated with TNF-alpha. The activation of promoter activity by TNF-alpha was abolished when a mutation was inserted into kappaB1 or kappaB2. These data indicated that pIgR gene expression induced by TNF-alpha is transcriptionally regulated via activation of NF-kappaB. In addition, there is a possibility that another factor may act in concert with NF-kappaB.

Binding Sites↗

PubMed2000

Cytokine profiles of T-lymphocytes from gingival tissues with pathological pocketing.

Periodontal disease is an infection in which destruction occurs at sites remote from the infection, resulting in pathological pocketing. Intervening between the infection and the destruction is a dense mononuclear inflammatory infiltrate. It has been suggested that this infiltrate might have characteristics and the destructive potential of Th1-type T lymphocytes. To ascertain the nature of the infiltrates we investigated the expression of mRNA for IL-2, IL-5, and IFN-gamma by gingival mononuclear cells (GMC) from healthy (n = 8) or adult periodontitis (AP) patients (n = 25) by using cytokine-specific reverse-transcription/polymerase-chain-reaction (RT-PCR). GMC, as obtained from patients' tissues, expressed IL-2, IFN-gamma, or IL-5 mRNA. Significantly higher proportions of GMC from AP patients expressed IL-2 and IFN-gamma mRNA than did those from healthy subjects. IFN-gamma was the most consistent cytokine message detected. In other experiments, gingival T-lymphocytes (n = 12) and CD4+ and CD8+ gingival T-lymphocytes (n = 16) were isolated from gingival tissues removed surgically from AP patients. AP gingival T-lymphocytes expressed mRNA for IL-2, IFN-gamma, or IL-6 prior to stimulation. After stimulation with Con A, the cells significantly up-regulated IL-5 and IL-6 message expression. Both CD4+ and CD8+ gingival T-lymphocytes expressed IFN-gamma, IL-5, and some IL-2. This cumulative cytokine profile observed in these experiments is consistent with the predominance of Th1-type cells in pathological tissues and with Th2-type cells, which can also be present, being up-regulated under appropriate stimulation. Importantly, CD4+ and CD8+ lymphocytes were shown to express T1- and T2-type cytokine message, emphasizing the potential for CD8+ T-lymphocytes to participate in periodontal disease pathology.

Adolescent↗

PubMed2000

A histochemical study of inflammatory lesions of the maxillary sinus mucosa using biotinylated lectins.

The distribution of glycoconjugates in normal maxillary sinus tissues, in cases of maxillary sinusitis and in postoperative maxillary cysts (POMC), was examined using seven different lectins as probes. The results showed that wheatgerm agglutinin (WGA), peanut agglutinin (PNA), Ulex europaeus agglutinin-1 (UEA-1), Ricinus communis agglutinin-1 (RCA-1), and concanavalin A (ConA) strongly react with the cilia and goblet cells. The binding of WGA, PNA, UEA-1, and RCA-1 was increased in maxillary sinusitis and POMC compared with normal maxillary sinus epithelium, whereas that of ConA was decreased. The decreased binding of ConA suggested that there were fewer mannoside residues in the maxillary sinus epithelium in the inflammatory lesion. The PNA bound to the cilia, goblet cells and mucous glandular cells in maxillary sinusitis and POMC, but not in normal, uninflamed cells, indicating that D-galactose was produced by the inflammatory condition. Similar binding patterns of PNA and RCA-1 were found in the cilia and on the surface of the epithelium and in the goblet cells. It is assumed that the carbohydrate moiety in the sinus mucosa is altered in inflammatory conditions.

Acetylgalactosamine↗

PubMed1999

Secretory IgA-mediated basophil activation. II. Roles of GTP-binding regulatory proteins and phosphatidylinositol 3-kinase.

Secretory IgA (sIgA) is the most abundant Ig isotype in mucous secretions in the upper and lower airways, where basophils exert effector functions during allergic inflammation. We recently demonstrated that immobilized sIgA on Sepharose beads is capable of inducing basophil degranulation ( approximately 15% of total histamine). To investigate the detailed mechanisms of this degranulation, we established in this study a new assay system for sIgA-mediated basophil activation. Immobilized sIgA on a plastic surface induced strong histamine release ( approximately 50% of total histamine) comparable to anti-IgE, and we analyzed the nature of basophil signal transduction by sIgA using various inhibitors. sIgA-induced basophil histamine release was inhibited completely by pertussis toxin, but anti-IgE-induced release was not affected. sIgA-mediated release was also inhibited by phosphatidylinositol 3-kinase (PI 3-kinase) inhibitor, wortmannin. These results strongly suggest that sIgA activates basophils via an IgE-independent novel mechanism involving both Gi protein and PI 3-kinase.

Basophils↗

PubMed1999

Predicting tumor metastasis in patients with oral cancer by means of the proliferation marker Ki67.

Recent developments of a Ki67 antibody to recombinant parts of the Ki67 nuclear antigen have provided a marker for tumor proliferation. In the present study, biopsy specimens were obtained from 20 patents with squamous cell carcinoma of the oral cavity at various sites, who also received a regional neck dissection. The patients' mean age was 61 years. Normal mucosa obtained from the surgical materials of 10 patients with a non-tumor condition was also examined as a control. The expression of Ki67 was examined immunohistochemically and the labeling index (LI) assessed in the biopsy specimens. The patients were divided into two groups; patient who already had a regional lymph node metastasis at the time of tumor resection and patient without any metastasis. All of the oral carcinoma and normal mucosa specimens were positive for Ki67, while the magnitude of staining showed a wide variation. The median LI of the patients with metastasis and without metastasis was 37.63 +/- 8.30 and 20.40 +/- 4.22 respectively, while the normal mucosa control was 7.62 +/- 1.70. The results of this study suggest that an immunohistochemical examination of the biopsy materials for the Ki67 antigen and assessed LI index should prove useful for the prediction of lymph node metastasis.

Adult↗

PubMed1998

Secretory IgA induces degranulation of IL-3-primed basophils.

We examined whether secretory IgA (sIgA), known to mediate eosinophil stimulation, has an effect on basophil functions. An immobilized preparation of sIgA, but not of monomeric IgA, induced histamine release (approximately 15% of total histamine contents) from human basophils in vitro. sIgA-induced basophil histamine release was totally dependent on pretreatment with IL-3. IL-5 and granulocyte-macrophage CSF also primed basophils for sIgA-mediated release. Exogenous divalent ions, i.e., Ca2+ and Mg2+, were essential for sIgA-mediated basophil degranulation, and the degranulation was completed within 45 min. A newly synthesized lipid mediator, leukotriene C4, was also liberated from IL-3-primed, sIgA-stimulated basophils. Enzyme digestion experiments revealed that the (Fc)2 x secretory component portion of sIgA is important for sIgA-mediated basophil activation, but the functional binding sites of sIgA on basophils were surmised to be different from FcalphaR. These observations reveal the novel finding that sIgA is able to stimulate basophils as well as eosinophils. Since sIgA is the most abundant Ig isotype in the secretions from mucosal tissues, and basophils are active participants in allergic late-phase reactions, sIgA-mediated basophil mediator release is potentially involved in exacerbation of the inflammation associated with allergic disorders.

Basophils↗

PubMed1998

Molecular maturation and functional expression of mouse polymeric immunoglobulin receptor.

Mouse polymeric immunoglobulin receptor (pIgR) cDNA was stably introduced into a hamster-derived fibroblastic cell line, Chinese hamster ovary (CHO) cell, by the calcium phosphate method. Surface expression of pIgR was detected by immunostaining and FACS analysis. The immunoprecipitated products of cell lysates revealed that the molecular mass of the most mature form of pIgR was approximately 120 kDa. Western blotting and metabolic labeling experiments followed by immunoprecipitation with an anti-mouse secretory component (SC) Ab demonstrated the existence of a 110 kDa immature form of pIgR. The reason for the existence of two forms of pIgR molecule was examined by conducting pulse-chase experiments which revealed the pIgR underwent molecular maturation. During this process, the 110 kDa form of pIgR was converted into a 120 kDa form by glycosylation. Moreover, tunicamycin treatment revealed the core form of pIgR had a molecular mass of approximately 100 kDa. The pIgR expressed on the surface of the transfectant could specifically bind and take up mouse polymeric IgA (MOPC 315), suggesting that, at least in this mouse system, cell type-specific molecules are not necessary for surface pIgR expression and polymeric immunoglobulin (pIg) binding and uptake.

Animals↗